Burkholderia pseudomallei, the etiological agent of melioidosis, is a Gram-negative, facultatively anaerobic, motile bacillus that is responsible for a broad spectrum of illnesses observed in both humans and animals. While epidemiological surveys have demonstrated that B. pseudomallei is endemic to regions that typically border the equator, the incidence of disease is particularly high in South-East Asia and Northern Australia. In north eastern Thailand alone, an estimated 20% of community acquired septicaemias and approximately 40% of deaths due to complications associated with bacterial sepsis can be attributed to this organism. The manifestations of melioidosis are commonly represented by acute, sub-acute and chronic illnesses, with the clinical manifestations often being mistaken for malaria, plague, pneumonia and miliary tuberculosis. Infections are typically acquired via inhalation or aspiration, ingestion or via the direct contact of damaged surface tissues with contaminated waters or soils. Burkholderia mallei, the etiological agent of glanders, is a Gram-negative bacterium that is responsible for disease in donkeys, mules, horses and occasionally humans. Unlike the environmental saprophyte B. pseudomallei, however, B. mallei does not persist in nature outside of its soliped hosts. While B. mallei and B. pseudomallei are genotypically similar, significant phenotypic differences do exist between the two pathogenic species. Although glanders is one of the oldest diseases known to man, relatively little is known about the pathogenesis of disease caused by B. mallei. This phenomenon is due primarily to the lack of disease in North America along with the fact that B. mallei can be a particularly dangerous organism to study even in a controlled laboratory environment. B. pseudomallei is known to resist the bactericidal activity of both reactive oxygen and nitrogen intermediates as well as to survive and multiply within several mouse and human macrophage cell lines (see electronmicrograph). In order to identify genetic loci associated with these virulence phenotypes, specifically those genes that are up or down-regulated in an intracellular environment, we use standard macrophage uptake assays, recover the intracellular bacteria and isolate bacterial mRNA to probe DNA micro-arrays. The data sets obtained from these studies are used to identify genes required for intracellular survival. Genes of interest are disrupted by allelic exchange and their function(s) are defined using both in vivo model systems (mouse macrophage invasion, infectivity in hamsters etc).